The present invention relates to glutathione derivatives and their dosage forms that permit an efficient expression of a hematopoiesis promoting activity in the target site of action of the bone marrow when they are administered in the living body for the purpose of promoting hematopoiesis. More specifically, it relates to methods of facilitating the stability in vivo, without affecting the biological activity, of xcex3-glutamyl S (benzyl) cysteinyl-R(xe2x88x92)-phenyl-glycine by providing the same with a long-chain alkyl or an alkenyl ester having 12 carbons or more, as well as of imparting a structure that is capable of being efficiently distributed in the target organ of bone marrow, and to methods of providing safe and effective dosage forms of colloidal compositions for injections that are an important dosage regimen.
The administration of chemotherapeutic agents in the treatment of cancer or the suppression of hematopoietic functions by radiotherapy can cause neutropenia and thrombocytopenia as side effects. In neutropenia among them, an immunological activity is decreased as compared to the normal healthy state so that there is an increased risk of being exposed to severe infections. Therefore, granulocyte colony-stimulating factor (G-CSF), a biologically active protein having an effect of promoting the recovery of neutrophil counts in the peripheral blood, has been clinically administered with satisfactory pharmacological results. However, since it has been produced in cultured cells using the gene recombinant technology, its high production cost is posing a problem from a medical economic viewpoint. In addition, the dosage regimen thereof has been established only for injections. Considering-the pains inflicted to patients and the simplicity of use, there is a need for the development-of drugs that can be administered by other dosage forms such as oral drugs in addition to injections.
Under the circumstances in which there is a need for low molecular weight hematopoiesis-promoting substances that can replace biologically active proteinaceous drugs such as G-CSF, that can be presented at low cost, and that can be administered via non-injection routes, xcex3-glutamyl ethyl ester S (benzyl) cysteinyl-R(xe2x88x92)-phenyl-glycyl ethyl ester (TER199) and its related compounds disclosed in the specification of WO9640205 were found to have an effect of promoting the in vitro colony formation of granulocytes and macrophage precursor cells. It was also confirmed that the compounds have an in vivo effect of promoting recovery of the number of cells such as neutrophils and platelets in the peripheral blood, using rat models in which the hematopoietic functions were suppressed by the administration of a chemotherapeutic agent (5-fluorouracil), although in an experiment where the dosage was as high as 60 to 120 mg/kg body weight and the method of administration (intraperitoneal administration) is not always clinically tolerated. Experiments carried out by the present inventors, however, have revealed that TER199 is very unstable in biological sample solutions such as rat blood and, hence, it is inefficient in exhibiting a hematopoiesis promoting activity. Furthermore, it was found that when high dosages such as were used in Examples of the specification of WO9640205 were administered intravenously for example, it can cause inflammation at the site of administration due to its physical properties such as low solubility and low tissue transfer. The foregoing suggested that TER199 disclosed in the specification of WO9640205 does not have the usefulness of being put into practical use based on the data.
Next, the WO9640205 specification describes that TER199 is an inhibitor of glutathione S-transferase (GST) having glutathione as the basic backbone structure. GST has been reported to be possibly involved in the resistance against anti-cancer agents and alkylating agents, and besides it is known that the effect of inhibiting the activity of GST can be found in, in addition to TER199, ethacrynic acid that is used as a diuretic, indomethacin, ketoprofen etc. that are used as non-steroidal anti-inflammatory drugs (Rinsho Kensa (Laboratory Tests), Vol. 39 (4), pp. 450-453 (1995)).
It is also very likely that the hematopoiesis-promoting activity in the bone marrow is related to the ability to inhibit GST. In fact, it has been reported that ethacrynic acid that has a structure entirely different from that of TER199 has an activity of inducing in vitro the differentiation of leukemia cells in cooperation with activated D3 (Leukemia Research, Vol. 20 (9), pp. 781-789 (1996)).
After intensive research to resolve the above problems associated with the conventional technology, the inventors of the present invention have found the following facts and have completed,the present invention.
Thus, the inventors were able to improve the stability in vivo of xcex3-glutamyl S (benzyl) cysteinyl-R(xe2x88x92)-phenyl-glycine (TER117) by providing the same with a long-chain alkyl or an alkenyl ester having 12 carbons or more, without affecting the biological activity related to the promotion of hematopoiesis, and to impart a structure that is capable of being efficiently. distributed in the target organ of bone marrow. In this case, consistent results were obtained by any of the following: a monoester in which either one of the carboxyl groups at the two sites in TER177 has been esterified; a diester in which one has been changed to a short- or middle-chain alkyl or alkenyl ester of 12 carbons or less and the other has been changed to a long-chain alkyl or alkenyl ester of 12 carbons or more; or a diester in which both have been changed to long-chain alkyl or alkenyl esters of 12 carbons or more. As used herein, chains having 1 to 2 carbons are defined as the short chains, chains having 3 to 11 carbons as the middle chains, and chains having 12 carbons or more as the long chains. By changing to long-chain esters, stability in vivo, more specifically stability against decomposition by esterase, was enhanced as compared to a diethyl ester TER199 disclosed as a preferred embodiment in WO9640205 specification. On the other hand, it was found in an in vitro experiment that the biological activity related to the promotion of hematopoiesis was equal to that of TER199. Furthermore, in long-chain esters having 12 carbons or more of TER117, the pharmacokinetic parameters, distribution volume and clearance, could be both decreased by using colloidal pharmaceutical compositions such as a fat emulsion, with a resultant discovery that the pharmacological effects can be efficiently exhibited in the target organ of bone marrow and the problem of safety related to the physical properties and physiological effects per se can be avoided.
Thus, the present invention relates to a glutathione derivative represented by the following formula (I): 
wherein, A represents H or a C1-C20 acyl group; R1 represents a C1-C26 alkyl group or a C3-C26 alkenyl group; and R2 represents H, a C1-C26 alkyl group or a C3-C26 alkenyl group, provided that the case wherein R1 represents a C1-C10 alkyl group or a C3-C10 alkenyl group and R2 represents H, a C1-C10 alkyl group or a C3-C10 alkenyl group is excluded, and a salt thereof.